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Circadian PER2 gene silencing suppresses lipid synthesis partly via inhibition of PPARG and SREBF1 in bovine mammary epithelial cells.

Y. F. Chen

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06-22-2020

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Abstract:

M76
Circadian PER2 gene silencing suppresses lipid synthesis partly via inhibition of PPARG and SREBF1 in bovine mammary epithelial cells.
Y. J. Jing1, Y. F. Chen*1, M. Z. Wang1, L. Y. Hu1, Q. Y. Xu1, Z. N. Xi1, J. J. Loor2. 1Yangzhou University Yangzhou, Jiangsu, China, 2University of Illinois Urbana, IL.

In non-ruminants it is well-established that biological rhythms play a profound role in coordinating whole-body metabolism. In dairy cows there is evidence that milk yield and milk fat content have rhythmic pattern thought to be regulated by circadian rhythms. The core circadian clock gene period 2 (PER2) is associated with mammary gland development and lipid synthesis in rodents, partly via regulating peroxisome proliferator-activated receptor gamma (PPARG). Whether such type of molecular link between circadian clock and lipid metabolism exists in bovine is unclear. We hypothesized that PER2 is associated with lipid metabolism in bovine mammary cells. To test this hypothesis, the bovine mammary tissue samples were obtained from 3 mid-lactation (averaged 110 d postpartum) cows and digested by collagenase to gain the primary bovine mammary epithelial cells (BMECs). Small interfering RNA (siRNA) technology was used to inhibit PER2 expression in primary BMECs. The primary BMECs were transfected with 3 siRNAs at 0, 12, 24, 36, 48, 60 h to screen out the best siRNA and its transfection time point. The lipid droplet was measured by red oil O staining, and the triacylglycerol (TAG) content of BMEC was determined with the tissue triglyceride assay kit (APPLYGEN, China). The lipid droplet and TAG content were determined at 36 h (36 h showed the greatest PER2 gene inhibitory effect of 84.7%) after the siRNA transfection. One-way ANOVA and Duncan's multiple comparison were used to conduct statistical analysis by SPSS software version 22.0 (statistical significance set at P < 0.05). Silencing of PER2 led to lower concentration of cellular lipid droplets and TAG levels in BMECs (P < 0.05). In addition, PER2 silencing downregulated mRNA of ACACA, FASN, LPIN1 and SCD (P < 0.05), indicating an overall inhibition of lipogenesis and desaturation. The downregulation of PPARG and SREBF1 in response to PER2 silencing underscore the importance of circadian clock signaling and transcriptional regulation of lipogenesis. Therefore, data suggest that PER2 participates in the coordination of mammary lipid metabolism and may be a component of the control of lipid droplet and TAG synthesis in mammary cells.

Keywords: PER2 silencing, mammary epithelial cell, lipid metabolism.